Project Summary: Mantle Cell Lymphoma (MCL) cells exhibit genetic alterations involving the cell cycle G1/S checkpoint- regulatory genes cyclin D1, CDKN2A, CDK4, as well as the cell growth and survival genes MYC and BCL2. MCL cells also display increased B cell receptor signaling and transcriptional activity of NFkB, making them sensitive to the lethal activity of the Bruton?s tyrosine kinase (BTK) inhibitor ibrutinib. Whereas treatment with ibrutinib induces clinical responses and improves survival, primary refractoriness or eventual emergence of resistance to ibrutinib is common, preventing durable remissions in MCL. Recently, mutations in CARD11/IKBKB/TRAF2/BIRC3/NIK, which lead to the activation of the classical or alternative NFkB signaling, have been documented, to confer resistance to ibrutinib. Our preliminary studies have demonstrated that treatment with the prototype BET (bromodomain and extra terminal) protein (BETP) antagonist (BETi) inhibits the mRNA and protein levels of the MCL-relevant oncogenes, including MYC, BCL-2 and CDK4/6. BETi treatment reduced the nuclear levels of NFkB, abrogated BRD4-dependent NFkB activity, and repressed its pro-growth and pro-survival target genes, including BTK, leading to apoptosis of human MCL cells. Notably, treatment with BETi alone also induced apoptosis of ibrutinib-resistant MCL cells. However, despite their promising anti-tumor activity, treatment with BETi leads to BRD4 protein accumulation, which limits BETi- mediated inhibition of the MCL-relevant oncoproteins including NFkB, thereby reducing the antitumor activity of BETi treatment. However, we have recently determined that treatment with the hetero-bifunctional BETP- PROTAC (Proteolysis Targeting Chimera), e.g., ARV-825 or ARV-771, which recruit BETPs to the E3 ubiquitin ligase cereblon or VHL, respectively, causes efficient, and prolonged degradation of BRD4, leading to inhibition of the MCL-relevant oncoproteins including NFkB. Based on this, we hypothesize that treatment with BETP- PROTAC will induce significantly more apoptosis than BETi, as well as exert synergistic lethality and anti- tumor efficacy with ibrutinib or with anti-BCL2 or CDK4/6 kinase inhibitor against cultured and patient-derived (PD) primary MCL cells. In AIM 1, we will determine the in vitro lethal activity and elucidate its predictive minimal signature of genetic-mutations and gene-expression perturbations due to the BETP-PROTACs (ARV- 825 and ARV-771) versus clinically-relevant BETis (OTX015 and ABBV-075), alone and in combination with ibrutinib in cultured cell lines as well as patient-derived (PD), genetically-profiled, primary MCL cells. In AIM 2, we will determine the combined in vitro lethal activity, and its predictive minimal gene-expression perturbations signature, due to treatment with the combinations of BET-PROTAC versus BETi with anti-BCL2 or anti-CDK4/6 inhibitor against the cultured and genetically-profiled, PD, ibrutinib-sensitive (IS) or ibrutinib-persister/resistant (IR) MCL cells. In AIM 3, we will determine the in vivo efficacy of co-treatment with ARV-771 or ABBV-075 and venetoclax or palbociclib, utilizing the cultured cell line and PD xenograft (PDX) models of IS or IR MCL cells.